We compared matched fibroblasts and 11 hiPSC clones analyzed at early, intermediate and late passages (Figure 5D–E, Figure S5B) using both the 27K and 450K DNA Methylation arrays (Table S5C–D). Shortly after reprogramming, there was an increase in XIST expression and in overall X chromosome DNA methylation. This was consistent with the higher level of X chromosome DNA methylation seen in a subset of female hPSC samples compared to the female somatic samples (X-Cluster 4 and X-Cluster 5; Figure 5B). At later passages, 8/11 hiPSC clones showed focal loss of XCI, indicated by loss of DNA methylation and increased mRNA expression, in the same regions observed in the hPSC collection as a whole (Figure S5B). The 3/11 hiPSC clones that retained full XCI at late passage also retained high levels of XIST expression (Figure S5B). There were two hiPSC clones (iPS3 and iPS7) that had intermediate levels of XIST expression at late passage (†, Figure S5B), but showed focal loss of XCI, consistent with the XIST threshold effect suggested above (Figure 5C–D). Using allele-specific RT-PCR, we confirmed that loss of
